Polynucleotide arrays (such as DNA or RNA arrays), are known and are used, for example, as diagnostic or screening tools. Such arrays include regions of usually different sequence polynucleotides arranged in a predetermined configuration on a substrate. These regions (sometimes referenced as “features”) are positioned at respective locations (“addresses”) on the substrate. In use, the arrays, when exposed to a sample, will exhibit an observed binding or hybridization pattern. This binding pattern can be detected upon interrogating the array. For example, all polynucleotide targets (for example, DNA) in the sample can be labeled with a suitable label (such as a fluorescent dye), and the fluorescence pattern on the array accurately observed following exposure to the sample. Assuming that the different sequence polynucleotides were correctly deposited in accordance with the predetermined configuration, then the observed binding pattern will be indicative of the presence and/or concentration of one or more polynucleotide components of the sample.
Biopolymer arrays can be fabricated by depositing previously obtained biopolymers (such as from synthesis or natural sources) onto a substrate, or by in situ synthesis methods. Methods of depositing obtained biopolymers include dispensing droplets to a substrate from dispensers such as pin or capillaries (such as described in U.S. Pat. No. 5,807,522) or such as pulse jets (such as a piezoelectric inkjet head, as described in PCT publications WO 95/25116 and WO 98/41531, and elsewhere). For in situ fabrication methods, multiple different reagent droplets are deposited from drop dispensers at a given target location in order to form the final feature (hence a probe of the feature is synthesized on the array stubstrate). The in situ fabrication methods include those described in U.S. Pat. No. 5,449,754 for synthesizing peptide arrays, and described in WO 98/41531 and the references cited therein for polynucleotides. The in situ method for fabricating a polynucleotide array typically follows, at each of the multiple different addresses at which features are to be formed, the same conventional iterative sequence used in forming polynucleotides from nucleoside reagents on a support by means of known chemistry. This iterative sequence is as follows: (a) coupling a selected nucleoside through a phosphite linkage to a functionalized support in the first iteration, or a nucleoside bound to the substrate (i.e. the nucleoside-modified substrate) in subsequent iterations; (b) optionally, but preferably, blocking unreacted hydroxyl groups on the substrate bound nucleoside; (c) oxidizing the phosphite linkage of step (a) to form a phosphate linkage; and (d) removing the protecting group (“deprotection”) from the now substrate bound nucleoside coupled in step (a), to generate a reactive site for the next cycle of these steps. The functionalized support (in the first cycle) or deprotected coupled nucleoside (in subsequent cycles) provides a substrate bound moiety with a linking group for forming the phosphite linkage with a next nucleoside to be coupled in step (a). Final deprotection of nucleoside bases can be accomplished using alkaline conditions such as ammonium hydroxide, in a known manner.
The foregoing chemistry of the synthesis of polynucleotides is described in detail, for example, in Caruthers, Science 230: 281-285, 1985; Itakura et al., Ann. Rev. Biochem. 53: 323-356; Hunkapillar et al., Nature 310: 105-110, 1984; and in “Synthesis of Oligonucleotide Derivatives in Design and Targeted Reaction of Oligonucleotide Derivatives”, CRC Press, Boca Raton, Fla., pages 100 et seq., U.S. Pat. No. 4,458,066, U.S. Pat. No. 4,500,707, U.S. Pat. No. 5,153,319, U.S. Pat. No. 5,869,643, EP 0294196, and elsewhere
Polynucleotide arrays have previously been provided in two formats. In one format, the array is provided as part of a package in which the array itself is disposed on a first side of a glass or other transparent substrate. This substrate is fixed (such as by adhesive) to a housing with the array facing the interior of a chamber formed between the substrate and housing. An inlet and outlet may be provided to introduce and remove sample and wash liquids to and from the chamber during use of the array. The entire package may then be inserted into a laser scanner, and the sample exposed array may be read through a second side of the substrate.
In another format, the array is present on an unmounted glass or other transparent slide substrate. This array is then exposed to a sample optionally using a temporary housing to form a chamber with the array substrate. The slide may then be placed in a laser scanner to read the exposed array. Most slide scanners require that the user manually insert the slide into a holder within the scanner. Some scanners allow the slide to rest on a surface while others clamp it to a known location using various types of guides. The present invention realizes that this technique creates a number of potential problems. First, since the array itself is unprotected it is subject to damage. Any damage is extremely undesirable for a number of reasons. For example, slight damage, such as fingerprints or scratches may occur to the sample exposed array which is not noticed. Such damage could lead to incorrect readings with serious consequences in interpretation of results. Also, it is not uncommon for the slides to be broken during insertion or removal from these scanners. Slide glass is easily chipped or broken. Losing a slide at this stage of the experiment can be extremely costly. Typically, the arrayed slides cost several hundred dollars and may involve long lead times. The samples under test may be from tumors or other hard-to-obtain sources. The fluorescent dyes typically employed are currently quite expensive. Therefore, a broken slide represents the loss of many hundreds of dollars and many hours of work. Thus, the present invention realizes that it is preferred to have a safer method and means of handling these slides. Furthermore, given that the individual features within the arrays on the surface of such slides are on the order of 10 to 120 microns in size and the importance of gathering all possible fluorescent signal, it is desirable to reference and hold these slides precisely. However, the present invention further realizes that precision placement usually involves firm surfaces and forcibly clamping the slides, which actions can result in slide breakage or array damage. If the slide is simply placed into a chamber to avoid clamping, large positional tolerances are needed which reduce the detection quality of the signals from the surface. Gathering all possible fluorescent signal from each feature on the array also requires that other sources of noise are minimized.
It would be desirable then to provide a means which could protect moieties, such as an exposed array, carried on a slide and protect the slide itself from breakage, which is relatively easy to use without requiring extensive manipulations of the slide, and which can aid in precisely positioning the slide (and hence the moieties) in a reader for reading of the exposed array.